The invention relates to a method for linking nucleic acids and/or glycosaminoglycans and/or glycosaminoglycan mimetics to polar/hydrophilic materials.
Nucleic acids such as deoxyribonucleic acid (DNA) and ribonucleic acid (RNA) and glycosaminoglycans (GAG) play a crucial part in cell functioning. DNA typically encodes proteins, RNA is typically an intermediate in protein synthesis and glycosaminoglycans play an important role in fundamental processes such as growth, differentiation and blood coagulation.
Nucleic acids such as DNA and RNA consist of covalently linked chains of deoxyribonucleotides (DNA) or covalently linked chains of ribonucleotides (RNA). Each nucleotide in DNA and RNA contains three characteristic components (1) a nitrogenous heterocyclic base which is a derivative of either pyrimidine or purine and (2) a pentose sugar and (3) a molecule of phosphoric acid that forms a negatively charged “backbone”.
Glycosaminoglycans are polysaccharide molecules made up of disaccharide repeating units containing a derivative of an amino sugar, either glucosamine or galactosamine wherein at least one of the sugars in a disaccharide unit has a negatively charged carboxylate or sulfate group. Glycosaminoglycans are heterogeneous in this respect and can contain 1 to 4 negative charges per disaccharide.
Nucleic acids and glycosaminoglycans can both be viewed as polymeric molecules with a “backbone” having a series of negative charges at neutral pH.
As a result of the development of molecular biological techniques, nucleic acids play an increasingly large part in the analysis of biological material. Fragments of DNA or RNA that are specific for example for causative organisms of disease (bacteria, viruses, etc.) can be used for detection of these organisms and diagnosis of hereditary diseases also occurs at nucleic acid level. Aside from the diagnostic importance of DNA/RNA, nucleic acids are also often employed for synthesis of specific proteins via recombinant DNA techniques and PCR (polymerase chain reaction).
Heparin has been demonstrated to have strong anti-coagulant properties and is useful in the production of medical devices where it may serve to prevent the binding of thrombocytes (blood platelets) to said medical devices and inhibit the formation of neointima formation (intimal hyperplasia).
Devices which are coated with nucleic acids and/or glycosaminoglycans for use in diagnostics or medicine should be able to bind a significant amount of nucleic acids and/or glycosaminoglycans in a tight association so that said nucleic acids and/or glycosaminoglycans are not significantly lost from the surfaces of devices coated with or linked to these macromolecules.
EP 797 778 relates to a method for preparing non-proteinaceous strongly negatively charged macrobiomolecules which are linked to plastic. Plastics may be considered as high molecular weight materials that contain synthetic or semisynthetic organic substances made by polymerization or condensation or derived from a natural material by chemical treatment that are molded, cast, extruded, drawn or laminated under various conditions. The method described in EP 797 778 is based on the finding that negatively charged molecules can be bound to apolar plastic surfaces like polystyrene using high salt conditions. The fact that the binding only occurs (is strongly increased) at very high salt concentrations strongly suggests that hydrophobic interactions are the basis of the bonding. The mechanism of operation is described in this patent application as resulting from the fact that the salt removes the water coat around the negatively charged molecules and shields the negative charges such that the interaction with apolar plastic, for example polystyrene, is strongly improved.
Hydrophobic interactions are of course expected to be much stronger to non-polar hydrophobic surfaces like plastics such as polystyrene than to polar/hydrophilic surfaces.
A number of materials used for diagnostic tests and medical devices are polar/hydrophilic materials. It should be reasoned therefore that binding of charged biomolecules such as nucleic acids and/or glycosaminoglycans to these polar/hydrophilic materials occurs through electrostatic interactions, and that promoting hydrophobic interactions would be of no advantage.
An object of the present invention is to provide a method for strongly linking nucleic acids and/or glycosaminoglycans to materials useful for diagnostic tests and assays and medical devices.
A further object of the present invention is to provide materials strongly linked to nucleic acids and/or glycosaminoglycans as well as devices comprising or consisting of materials strongly linked to nucleic acids and/or glycosaminoglycans